The Syrian golden hamster is the classic animal model for the vivo study of respiratory carcinogenesis. Hamster tracheal explant cultures have been used to study in vitro carcinogenesis. Because of the advantages of using serum-free media in carcinogenesis studies in vitro, various formulations of media were tested for the explant and subsequent cell culture of hamster respiratory epithelia. Media CMRL-1066, 199, MCDB 152, and modified MEM were tested. All media had growth factors replacing fetal bovine serum. Optimal outgrowths of epithelial cells from tracheal rings were obtained in CMRL-1066 and modified MEM. However, more cells underwent terminal differentiation in medium CMRL-1066 due to its high calcium concentration than in the modified (low calcium) MEM containing insulin, hydrocortisone, bovine pituitary extract and epidermal growth factor. Additional tests are under way on media which allow the replicative culture of cells from the outgrowths. It was found that tracheal rings can be cryopreserved at -70 C for extended periods; this procedure has greatly facilitated the use of tracheal rings in these studies. Similar techniques were developed for the explant culture of hamster larynx and bronchi in serum-free medium. Suties comparing response of the various segments of the hamster respiratory tract are ongoing, using treatments with carcinogens having different specificity in different respiratory tract segments. Tracheal rings removed from hamsters pretreated with diethynitrosamine were grown in explant culture in serum-free CMRL-1066 and were compared to non-treated controls; preliminary results suggest that carcinogen treatment in vivo appears to facilitate the outgrowth of cells from the tracheal rings. A qualitative response to the in vitro treatment of tracheal rings with 3-methylindole and 2-methylnaphthalene has been observed by comparing the extent of outgrowth of cells from the treated rings to untreated controls.